This U54 application describes the Northwest Genome Engineering Consortium, an investigative team which will pursue an interdisciplinary approach to develop concepts, methods, and tools for gene repair, a new approach to the treatment of monogenic disorders of the bone marrow and lymphoid systems. Despite advances in conventional and 2nd generation therapies, monogenic disorders of lymphohematopoiesis have significant health impacts both within the US and on a global scale. Thus, these diseases are an important area for the development of new treatment approaches. Genome engineering technology has progressed to the point that it is presently feasible to modify individual defective genes in primary human cells, a process termed gene repair. The goal of the ^orthwest Genome Engineering Consortium (NGEC) is to develop and apply gene repair to the treatment of monogenic lymphohematopoietic disorders. This goal will be accomplished through two major scientific aims. The first aim will support a team approach that combines computational protein design, directed protein evolution, structural analysis, and in vitro biochemical and molecular analyses with the goal of pioneering methods for producing artificial LAGLIDADG homing endonucleases (LHEs), for use in a variety of types of genome engineering applications. The second aim will support a team approach to the application of LHE's for site specific gene repair in hematopoietic cells. This approach will combine novel LHE's created in Aim 1 with the use of non-integrating lentiviral vectors for the introduction of an LHE and a repair/modification template to hematopoietic stem cells for the purpose of gene repair. It will also explore new methods for manipulating endogenous DMA repair mechanisms to enhance the in situ efficacy of LHE-induced gene repair in hematopoietic stem cells, and develop methods for autologous hematopoietic stem cell engraftment optimized for gene repair applications. In addition to its focus on applying gene repair to hematopoietic stem cells, the NGEC will contribute to the development of genome engineering as a broader scientific discipline through: 1) a post-doctoral training program which will provide an opportunity for qualified fellows to obtain experience and training in the process of LHE-based genome engineering; and 2) an annual Workshop on Genome Engineering, which will be open to students and faculty from any institution, and will include talks by prominent outside investigators working in genome engineering related fields.